Portable interferometric device

ABSTRACT

The present invention provides a novel simple, portable, compact and inexpensive approach for interferometric optical thickness measurements that can be easily incorporated into an existing microscope (or other imaging systems) with existing cameras. According to the invention, the interferometric device provides a substantially stable, easy to align common path interferometric geometry, while eliminating a need for controllably changing the optical path of the beam. To this end, the inexpensive and easy to align interferometric device of the invention is configured such that it applies the principles of the interferometric measurements to a sample beam only, being a single input into the interferometric device.

REFERENCES

The following is a list of publications that might be pertinent forunderstanding the background of the technology to which the inventionrelates:

-   1. N. T. Shaked, Y. Zhu, N. Badie, N. Bursac, and A. Wax, J. of    Biomed. Opt. Lett. 15, 030503 (2010).-   2. G. Popescu, T. Ikeda, R. R. Dasari, and M. S. Feld, Opt. Lett.    31, 775 (2006).-   3. H. Ding and G. Popescu, Opt. Express 18, 1569 (2010).-   4. J. Jang, C. Y. Bae, J.-K. Park, and J. C. Ye, Opt. Lett. 35, 514    (2010).-   5. B. Kemper, A. Vollmer, C. E. Rommel, J. Schnekenburger, and G.    von Bally, J. Biomed. Opt. 16, 026014 (2011).-   6. N. T. Shaked, L. L. Satterwhite, G. A. Truskey, M. J. Telen,    and A. Wax, “Quantitative microscopy and nanoscopy of sickle red    blood cells performed by wide field digital interferometry,” J.    Biomed. Opt. 16, 030506 (2011).-   7. V. Mico, Z. Zalevsky, and J. Garcia, “Common-path phase-shifting    digital holographic microscopy: a way to quantitative phase imaging    and superresolution,” Opt. Commun. 281, 4273-4281 (2008).-   8. V. Micó and J. Garcia, “Common-path phase-shifting lensless    holographic microscopy,” Opt. Lett. 35, 3919-3921 (2010).-   9. P. Bon, G. Maucort, B. Wattellier, and S. Monneret, “Quadriwave    lateral shearing interferometry for quantitative phase microscopy of    living cells,” Opt. Express 17, 13080-13094 (2009).-   10. M. Lee, O. Yaglidere, and A. Ozcan, “Field-portable reflection    and transmission microscopy based on lensless holography,” Biomed.    Opt. Express 2, 2721-2730 (2011).-   11. P. Kolman and R. Chmelík, “Coherence-controlled holographic    microscope,” Opt. Express 18, 21990-22003 (2010).-   12. Z. Monemhaghdoust, F. Montfort, Y. Emery, C. Depeursinge, and C.    Moser, “Dual wavelength full field imaging in low coherence digital    holographic microscopy,” Opt. Express 19, 24005-24022 (2011).-   13. Z. Wang, L. J. Millet, M. Mir, H. Ding, S. Unarunotai, J. A.    Rogers, M. U. Gillette, and G. Popescu, “Spatial light interference    microscopy (SLIM),” Opt. Express 19, 1016-1026 (2011).-   14. N. T. Shaked, T. M. Newpher, M. D. Ehlers, and A. Wax, “Parallel    on-axis holographic phase microscopy of biological cells and    unicellular microorganism dynamics,” App. Opt. 49, 2872-2878 (2010).

FIELD AND BACKGROUND

This invention is generally in the field of interferometry, and relatesto a system and method for interferometric measurements used forinspecting samples. The invention can be particularly used with amicroscope or other imaging systems to acquire quantitative inspectionof transparent, semi-transparent or reflective samples.

Interferometric microscopy, also known as wide-field interferometricphase microscopy (IPM) or digital holographic microscopy can be used tosimultaneously record the quantitative spatial profiles of both theamplitude and the phase of the measured samples. Using interferometricmicroscopy, time recording of the phase profile can yield remarkableoptical thickness or optical-path-delay stability of less than ananometer, with acquisition rates of several thousands of full framesper second, and without the need for using contrast agents such asflorescence dyes. As the technique provides the optical thickness pereach spatial point on the sample, various relevant morphological andmechanical parameters of the sample can be obtained in a non-contact,label-free manner IPM can be utilized for a wide range of applicationsincluding biological cells investigations, surface measurements,biometry, and others. IPM uses interference to record the complex wavefront (amplitude and phase) of the light interacted with the sample. Forbiological and medical applications, the ability to record the samplequantitative phase enables the user to see cells and organisms, whichare otherwise transparent due to the cell low absorption and scatteringof the transmitted light.

These unique advantages could be attractive for many clinicalapplications, so many IPM setups were presented over the years, and theycan be divided into various groups, such as setups that use common-pathgeometry [7-9] or separated reference and sample beam geometry [10],setups that use high-coherence source [6] or low-coherence source[11-13], setups that use on-axis (inline) geometry or off-axis geometry[4,5, 10-14]. However, not many options are available for commercialinterferometric microscopes compared to other microscopy techniques, andthis tool is mostly used by optical and biomedical engineers forresearch purposes. One reason for this is the difficulty to obtainhigh-quality and stable interference pattern with modest and portableequipment and without the need for an expert user. The commonly-usedinterferometric setups are usually constructed in open and custom-builtmicroscopes and operated by a user with knowledge in optics. To ensurethe stability of the interference pattern, the entire system ispositioned on an optical table to avoid mechanical vibrations and isboxed inside an enclosure to avoid differential air perturbationsbetween the interferometric arms.

Techniques aimed at or enabling higher stability of the interferencepattern with compact and portable designs have been developed. One ofthese systems is the interferometric chamber (InCh) microscope [1]. Inthis system, all the interferometric elements are encapsulated into asingle, rigid and factory-designed reflective chamber. Although thissystem uses common-path geometry (and thus can operate without anoptical table), it can still create off-axis interferograms of thesample (and thus only one frame is required for acquiring the amplitudeand the phase profiles of the sample, which is suitable for highlydynamic samples). However, the InCh microscope cannot use highmagnifications due to the fact that the microscope objective needs tocollect the tilted reference beam. In addition, this microscope requireshighly-coherent illumination sources since the optical path differencebetween the reference and the sample beams are twice the opticalthickness of the chamber. A similar technique is described in US2011/0242543. The system includes a light source for generating anillumination beam that propagates towards a sample. A sample holder mayhold the sample and include a partially reflective cover for allowing afirst portion of the illumination beam to pass therethrough to interactwith the sample to produce a sample beam that propagates substantiallyalong an optical axis. The cover may be oriented at an angle forreflecting a second portion of the illumination beam to produce areference beam that propagates at a predetermined angle with respect tothe optical axis. An imaging module may redirect the reference beamtowards the optical axis at a detection plane. A detector may interceptthe sample and reference beams and may generate a holographicrepresentation of the sample based on the beams.

Other setups for common-path or self-interference interferometry havebeen presented [2-5]. In one type of setups, a diffraction grating orother specialized optical elements are used, whereas in another type ofsetups, a Michelson interferometer in the output of a microscope isused, so that the sample beam interferes with itself, with thelimitation that half of the sample has to be empty. Many of the knowninterferometric setups, however, have the same main drawbacks ofbulkiness, non-portability and the requirement for specific opticalskills to align and use them. These shortcomings cause this technologyto largely remain in optical research laboratories, and thus it is notvery common in the industry or in clinics.

GENERAL DESCRIPTION

There is a need in the art for a novel interferometric device whicheliminates or at least significantly reduces the sensitivity of theinterferometric measurements to the effects of environmental conditions,such as mechanical vibrations and air perturbations to which theinterferometer is typically exposed. Also, there is a need in the fieldof interferometric measurements to increase the sensitivity of themeasurements by using low-coherence light sources. In addition, there isa need in the art for providing a portable and inexpensiveinterferometric module for microscopes or other imaging systems that canbe connected in their output, turning them to powerful interferometricimaging systems with the modest equipment and without the need of opticsknowledge, which will provide, for example, a device of acquiring livebiological cells in a label-free manner with sub-nanometric thicknessprecision or quality testing of elements with nanometer scalethicknesses after their production, or even assuring their production(for example, for controlling a lithography process).

The present invention provides a novel approach for interferometricmeasurements that solves the above problems and can be easilyincorporated in a microscope (or other imaging systems). According tothe invention, the interferometric device provides a substantiallycommon path for the sample and reference beams while eliminating a needfor controllably changing the optical path of the reference beam. Tothis end, the interferometric device of the invention is configured suchthat it applies the principles of the interferometric measurements to a“sample” beam only, being a single input into the interferometricdevice. In other words, in the invention, a beam that is split intosample and reference beams is the beam that has interacted with asample. Such input beam into the interferometric device of the inventionmay be provided as the output of a regular microscope, usuallypropagating towards the detector of the microscope. Thus, theinterferometric device of the invention may be installed in an opticalpath of light propagating from the sample through the optical systemdetector. The sampled beam, after its interaction with the sample isamplitude and phase modulated by the sample and then the beam propagatesinto the interferometric device, where it is split into a pair of beamsand combined again. Then, the beams propagate to the light detector(such as a microscope camera) where they interact and produce theinterference pattern, while one of these beams, on its way to thedetector, undergoes optical processing to remove the sample modulationand which beam will thus present a reference beam. This opticalprocessing is spatial filtering and is implemented by the passage of thebeam through two lenses and being reflected by a mirror located behind apinhole. This mirror-pinhole construction is located in a Fourier planewith respect to the sample plane of the imaging system. The beaminteracting with a pinhole in the Fourier plane results in filtering outof all the non-zero spatial frequencies of the beam. This configurationin which the mirror is located behind a pinhole enables to provide ahigh stability and a low noise. This configuration also enables tosimplify the alignment of the interferometer.

As described above, the interferometric device of the present inventioncan be attached to the camera port of a conventional microscope or otherimaging systems, while still obtaining high-quantity interferograms ofthe samples without the need for special optical skills or complicatedalignment prior to the experiment. Since the light splits and mergesonly after the output of the microscope, while using spatial filteringto erase the information from one of the beams before it merges with theother beam, this setup can be considered as a common-pathinterferometer, and thus it is more stable compared to regularinterferometers. In addition, this setup can be easily adjusted with alow-coherence light source, which creates clearer images with lesscoherent noise. In contrast to regular interferometers, since there isonly one sample beam till the output of the microscope, the user doesnot need to take into consideration the thickness of the constant sampleelements such as the cover-slips in order to create beam path matchingfor achieving interference with a low-coherence source. Due to theseadvantages, this setup achieved a low temporal and spatial noise levelsin the sub-nanometric range.

Thus, according to one broad aspect of the invention, there is providedan interferometric device comprising a light-directing opticalarrangement for directing light to an optical detector, wherein thelight directing optical arrangement is configured for defining first andsecond substantially overlapping optical paths towards the opticaldetector, the light directing optical arrangement comprises: a beamsplitter/combiner unit for receiving an input beam of the amplitude andphase modulation and splitting the input beams into first and secondlight beams, a first and second reflective surfaces accommodated in thefirst and second optical paths of the first and second light beams splitby the beam splitter/combiner to thereby direct the first and secondlight beams back to the beam splitter/combiner that directs the combinedbeam to the detector; a spatial filter comprises, a pinhole accommodatedin front of at one of the first and second reflective surfaces and aFourier optics assembly comprising two lenses, one being in a 4fconfiguration with respect to each other, the light directing opticalarrangement directing first and second optical beams along the first andsecond paths thereby enabling interaction between the first and secondbeams at the optical detector, an interference pattern resulting fromthe interaction of the first and second optical beams being therebyindicative of the amplitude and phase modulation, the beamsplitter/combiner unit directing one of the first and second light beamsthrough the spatial filter to enable amplitude and phase demodulationthereof and formation of a reference beam with respect to the othermodulated beam.

The pinhole is located in a predetermined Fourier plane (e.g. that ofthe surface being imaged).

In some embodiments, one of the first and second reflective surfacescomprises a retro-reflector. The retro-reflector may be built out of twomirrors connected in a right angle, which tilts sample beam, so that anoff-axis angle can be created between the sample and reference beams. Byusing this configuration, the interferometer of the present invention isconfigured to operate in off-axis geometry. It should be noted that theon-axis geometry might limit the capture of dynamic changes as more thana single exposure is needed to obtain the quantitative phase profile ofthe sample [12], while the sample might change between the frames ofacquisitions. In addition, complex elements, such as phase modulators,may be used to create the needed different exposures. The presentinvention also provides an off-axis interferometer operating in fulloff-axis geometry, enabling to obtain an interference on a large fieldof view (limited only by the coherence length of the source) and usingthe full frame rate of the camera, while still retaining the advantagesof the device portability, low cost, easy construction and alignment,even with a low-coherence source.

The off-axis configuration of the interferometer of the presentinvention presents a new simple-to-align, highly-portableinterferometer, which is able to capture wide-field, off-axisinterference patterns from transparent samples under low-coherenceillumination. This small-dimension and low-cost device can be connectedto the output of any microscope illuminated by a low-coherence sourceand yield sub-nanometric optical thickness measurements in a label-freemanner. The interference fringes have high spatial frequency, and theinterference area is limited only by the coherence length of the source,and thus it enables to obtain high-resolution quantitative images ofstatic and dynamic samples.

In some embodiments, the beam splitter/combiner unit comprises a cubebeam splitter.

In some embodiments, the first and second reflective surfaces are placedright at the outputs of the beam splitter/combiner unit.

In some embodiments, one lens is located at a distance equals to itsfocal length from the image plane of the imaging system.

In some embodiments, one of the reflective surfaces is located after oneof the lenses at a distance of the focal length of the lens.

In some embodiments, the detector is located at a distance of the focallength of one of the lens.

In some embodiments, the interferometer device comprises a phaseshifting device into one of the beam paths.

According to yet another broad aspect of the invention, there isprovided an optical system comprising: a beam splitter/combiner unit forreceiving an input beam of certain amplitude and phase modulation andsplitting the input beam into first and second light beams of the sameamplitude and phase modulation and combining reflections of the firstand second light beams to produce an output combined beam; a first andsecond reflective surfaces accommodated in the first and second opticalpaths of the first and second light beams to thereby direct the firstand second light beams back to the beam splitter/combiner that directsthe combined to the detector; a spatial filter comprising a pinholeaccommodated in front of the first mirror in the optical path of thefirst split light beam to apply amplitude and phase demodulation theretoand thereby form a demodulated reference beam with respect to the secondmodulated beam and a Fourier optics assembly comprising two lenses, onebeing in a 4f configuration with respect to each other; thereby enablingan interference pattern resulting from interaction of the reference andmodulated beams to be indicative of the amplitude and phase modulation.

According to yet further broad aspect of the invention, there isprovided a sample inspection system, comprising: light collecting andfocusing optics configured and operable for collecting an input beamfrom a predetermined sample surface and focusing it onto an image plane;an interferometer unit accommodated in a path of the light collected bythe light collecting and focusing optics, the intereferometer unitcomprising: a beam splitter/combiner unit for receiving the input beamof certain amplitude and phase modulation and splitting the input beaminto first and second light beams of the same amplitude and phasemodulation and combining reflections of the first and second light beamsto produce an output combined beam; a first and second reflectivesurfaces accommodated in the first and second optical paths of the firstand second light beams to thereby direct the first and second lightbeams back to the beam splitter/combiner that directs the combined tothe detector; a spatial filter comprising a pinhole accommodated infront of the first mirror in the optical path of the first split lightbeam being located in a Fourier plane with respect to the predeterminedsurface to thereby apply amplitude and phase demodulation thereto andform a demodulated reference beam with respect to the second modulatedbeam and a Fourier optics assembly comprising two lenses, and a secondlens being positioned in an 4 f configuration with the first lens; aninterference pattern resulting from interaction of the reference andmodulated beams in the image plane being thereby indicative of theamplitude and phase modulation.

BRIEF DESCRIPTION OF THE DRAWINGS

In order to understand the disclosure and to see how it may be carriedout in practice, embodiments will now be described, by way ofnon-limiting example only, with reference to the accompanying drawings,in which:

FIGS. 1A-1B are schematic figures of some possible configurations of theinterferometer of the present invention, positioned in the camera portof a regular microscope;

FIG. 2 is a plot showing comparative results about the temporal opticalpath delay between the interferometer of the present invention and aconventional Michelson interferometer;

FIG. 3 is an image of a quantitative thickness profile of a live redblood cell acquired with the interferometer of the present invention;

FIG. 4A is a schematic figure representing another possibleconfiguration of the interferometer of the present invention, accordingto another embodiment of the present invention;

FIG. 4B is a ray tracing of the sample and the reference beams as itwould be seen if they were on the same optical axis according to theembodiment of FIG. 4A;

FIG. 5 is a schematic figure of the interferometer of the embodiment ofFIG. 4A connected at the output of a regular microscope and illuminatedby a tunable low-coherence source;

FIGS. 6A-6B are spatial and temporal optical-path-delay profiles (OPDs)in a dry sample (OPDs) respectively; FIG. 6A represents an OPD standarddeviation across an image for 150 OPD images (spatial sensitivity) andFIG. 6B represents an OPD standard deviation between 150 OPD images oneach diffraction-limited spot (temporal sensitivity);

FIGS. 7A-7B are optical-path-delay or optical thickness maps of a volumephase holographic grating obtained under low-coherence illumination bythe off-axis interferometer of the present invention (FIG. 7A); and aMach-Zehnder interferometer (FIG. 7B) respectively;

FIG. 8 is a scanning electron microscope (SEM) image of a firstlithographed phase target;

FIGS. 9A-9C are optical-path-delay maps of a first phase target createdby Focused ion beam (FIB) lithography, containing variable depthselements, as obtained using: the off-axis interferometer of the presentinvention with a low-coherence source (FIG. 9A); an off-axisMach-Zehnder interferometer with a low-coherence source (FIG. 9B); andan off-axis Mach-Zehnder interferometer with a high-coherence source(HeNe laser) (FIG. 9C) respectively;

FIGS. 10A-10C are optical-path-delay maps of a second phase targetcreated by FIB lithography, containing variable depths elements, asobtained using: the off-axis interferometer of the present inventionwith a low-coherence source (FIG. 10A); an off-axis Mach-Zehnderinterferometer with a low-coherence source (FIG. 10B); and an off-axisMach-Zehnder interferometer with a high-coherence source (HeNe laser)(FIG. 10C) respectively;

FIGS. 11A-11B are red blood cell (RBC) optical-path-delay and physicalthickness maps obtained using a low-coherence source: the off-axisinterferometer of the present invention (FIG. 11A); an off-axisMach-Zehnder interferometer (FIG. 11B) respectively;

FIGS. 11C-11D are standard deviation of the optical-path-delay and ofthe physical thickness maps for: the off-axis interferometer of thepresent invention (FIG. 11C); and an off-axis Mach-Zehnderinterferometer (FIG. 11D) respectively; and;

FIG. 12 are measurements of Blepharisma organism swimming in water usingthe off-axis interferometer of the present invention and demonstratingthe device capabilities for quantitative imaging of fast dynamics onrelatively large field of view due to its true off-axis configuration.

DETAILED DESCRIPTION OF EMBODIMENTS

FIG. 1A presents a system 10 including an interferometric device 14which in the present not limiting example is incorporated in amicroscope being ported into the microscope output (replacing a digitalcamera typically installed there in the microscope). This configurationenables to connect a regular camera at the output of the device of thepresent invention. A magnified image of a sample from the microscope isformed by light 13 presenting amplitude and phase modulation of an inputlight incident on the sample, the amplitude and phase modulation beingindicative of the sample's effect on light passing therethrough. Theinterferometer device 14 comprises a light directing optical arrangementfor receiving input light 13 of certain amplitude and phase modulationand direct the light to an optical detector (e.g. digital camera) wherean interference pattern is detected being indicative of the amplitudeand phase modulation. The light directing optical arrangement of theinvention defines first and second substantially overlapping opticalpaths OP₁ and OP₂ towards the detector. These optical paths serve forpropagation of first and second optical beams of substantially the sameamplitude and phase modulation to thereby enable interaction betweenthese beams at the detector to produce the interference pattern. Thelight directing optical arrangement 14 includes a beam splitter/combinerunit BS for receiving input beam 13 of the amplitude and phasemodulation and splitting it into first and second light beams 13 a and13 b, and directing one of them (beam 13 b in the present example)through a spatial filter SP placed in the Fourier plane of one of theinterferometric arms to enable amplitude and phase demodulation thereofand formation therefrom a reference beam with respect to the othermodulated beam.

Further provided in the interferometric device 14 is a first and secondreflective surfaces M1 and M2 accommodated in the first and secondoptical paths of the first and second light beams to direct the firstand second light beams back to the beam splitter/combiner unit BS thatdirects the combined beam to the detector. The spatial filter SP isaccommodated in front of the second mirror M2.

Further provided in the interferometric device 14 is a Fourier opticsassembly configured for applying Fourier transform to an optical fieldof the input beam 13 and for applying inverse Fourier transform to anoptical field of a combined beam 15 propagating from the beam/splittercombiner to the detector. This Fourier optics assembly is thus formed bylenses L₁ and L₂, where lens L₁ is located at a distance equals to itsfocal length from the image plane of the imaging system. Thus, the imageplane in the output of the microscope is Fourier transformed by lens L1and then splits it into first and second beams by a cube beamsplitter/combiner BS. One of the beams (defined as the sample beam) isreflected by the element M1, located after lens L1 at a distance of thefocal length of lens L1, and then Fourier transformed back to the cameraplane using lens L2, located at a distance of the focal length of L2from M1, and the camera is located after L2 at a distance of the focallength of L2. The other beam (defined as the reference beam) isspatially filtered by a spatial filter SP constituted by pinhole,reflected by mirror M2 located right after the pinhole, and Fouriertransformed back to the camera plane by lens L2. The pinhole demodulatesone of the light beam thereby erasing the sample information by onlypassing the zero frequencies of the image Fourier transform, thuseffectively creating a reference beam with respect to the second beam,still containing the full sample information. Thus, the spatialfiltering effectively creates a reference beam by erasing the sampleinformation from one of the beams, and also increases the beam spatialcoherence and enables quantitative interference on the camera. Thedashed lines are directly transmitted light while the solid lines arethe image forming beams. The two beams are then reflected by element M1and mirror M2 and combined by the beam splitter/combiner. Another lensL2, positioned in 4f configuration with the first lens L2, back Fouriertransforms the combined beam and projects it onto a detector, e.g. adigital camera, where an interference pattern results from interactionof the reference and modulated beams in the image plane and aninterferogram of the sample is created.

By using this configuration, the two beams are on the same optical axis,causing the beams to propagate in the same direction after L2 lens. Theangle between the two beams is negligible and this causes an on-axisinterference pattern on the digital camera. Several phase-shiftedinterferograms would be required for the reconstruction process, whichcan be obtained by adding a phase shifting device into one of the beampaths. To reconstruct the sample profile using one interferogram, onecan shift the camera to the edge of the interference pattern so that theoff-axis interferogram appears on a small area where the fringes areparallel straight lines. However, this can be obtained in a very limitedfield of view, and thus the sample size that can be interferometricallyrecorded is significantly reduced.

In some embodiments, element M1 is a two-mirror construction such as aretro-reflector RR providing a novel interferometer having an off-axisconfiguration. This set-up will be described in detail further belowwith respect to FIG. 4A. The retro-reflector RR may comprise a cornerreflector, a cat's eye or a phase-conjugate mirror.

FIG. 1B presents a system 100 configured according to some embodimentsof the present invention, including an interferometric device 104 whichin the present not limiting example is incorporated in an microscopebeing ported into the microscope output (replacing a digital cameratypically installed there in the microscope). The microscope includes alight source 101, such as a low-coherence laser, a sample holder S, anda microscope objective MO. Also, the microscope may include a lightdirecting optics, such as a light deflector (mirror) M1 that directslight 102 from the light source onto a sample S, and a tube lens L₀. Theobjective lens and the tube lens create an appropriately magnified imageof the illuminated spot of the sample on an image plane located in theinterferometric device 104. The dashed lines are directly transmittedlight while the solid lines are the image forming beams.

The interferometer 104 receives the magnified image of the sample S fromthe microscope. This image is formed by light 103 presenting amplitudeand phase modulation of the input light 102 incident on the sample, theamplitude and phase modulation being indicative of the sample's effecton light passing therethrough. The interferometer 104 is configuredaccording to the invention as a light directing optical arrangement forreceiving input light 103 of certain amplitude and phase modulation anddirect to an optical detector (e.g. digital camera) where aninterference pattern is detected being indicative of the amplitude andphase modulation. The light directing optical arrangement of theinvention defines first and second substantially overlapping opticalpaths OP₁ and OP₂ towards the detector, and comprises a spatial filteraccommodated in one of the first and second optical paths.

The light directing optical arrangement 104 includes a beamsplitter/combiner unit BS for receiving input beam 103 of the amplitudeand phase modulation and splitting it into first and second light beams103 a and 103 b, and directing one of them (beam 103 b in the presentexample) through a spatial filter SP to enable amplitude and phasedemodulation thereof and formation therefrom a reference beam withrespect to the other modulated beam. Further provided in theinterferometric device 104 is a Fourier optics assembly configured forapplying Fourier transform to an optical field of the input beam 103 andfor applying inverse Fourier transform to an optical field of a combinedbeam 105 propagating from the beam/splitter combiner to the detector.This Fourier optics assembly is thus formed by lenses L₁ and L₂, wherelens L₁ is located in the image plane of the sample (i.e. the planebeing imaged).

Thus, device 104 receives input amplitude and phase modulated beam 103,Fourier transforms it by lens L₁ and then splits it into first andsecond beams by a cube beam splitter/combiner BS. The two beams are thenreflected by mirrors M and combined by the beam splitter/combiner. Thesetup provides an on-axis interferometric microscope, and an electriccontrol connected to one of the mirrors can create several phase shiftedinterferograms that are needed to retrieve the quantitative phaseprofile of the sample. However, to enable single-exposure operation,off-axis interferograms can be acquired by shifting the mirrors M or thecamera to high-spatial-frequency region, within the source coherencelength.

The configuration uses simple optical elements only and no gratings orother diffractive elements are used inside the interferometer 104. Itshould be understood that in most IPM setups, the beam is split to thereference and sample beam before interacting with the sample and thenthe beams propagate through different areas with different environmentalnoises. In contrast, in the present invention, the beam is split afterthe sample interaction and therefore provides an interferometer havingcommon-path geometry, where higher stability and lower noise isobtained. It should be noted that the input beam of the sample onlysplits in the end of the device, accordingly the proposed setup can beconsidered as a common-path interferometer, and its stability will besignificantly higher compared to regular interferometers. Moreover,since splitting the beam is done in the middle of the 4f device(coincides with the center of the beam splitter/combiner), theinterferometer 104 is closer to common path than other configurations inwhich the splitting is done in the beginning of the 4f device.Additionally, since the first and second beams, i.e. reference andsample beams, pass mostly through the glass of the cube beamsplitter/combiner, there are less differential air perturbations betweenthe interferometric arms, even if the interferometer is not boxed. Itshould also be noted that the mirrors in the interferometer 104 areplaced right in the outputs of the beam splitter BS and since the beamsare tightly focused on each of the mirrors M, it is significantly easierto match the beam paths, making it possible to obtain interference withlow-coherence sources.

According to another possible embodiment of the present invention, notshown in the figures, if a microscope with condenser annulus isavailable (such as in phase contrast microscope), the interferometer 104can use for the spatial filter a ring aperture instead of the pinhole Pin front of one of the mirrors M.

In order to demonstrate the capabilities of the invention, the inventorperformed an experiment with a device that is similar to the deviceillustrated in FIG. 1B with the following specifications: A temporallylow-coherence plane wave was created by passing a supercontinuumfiber-laser light (from SC400-4, Fianium) through a computer-controlledacousto-optics tunable filter (SC-AOTF, Fianium), selecting a centralwavelength of 633 nm with a full-width-at-half-maximum bandwidth of 6.6nm, as measured by a compact spectrometer (USB400, Ocean Optics). Thislow-coherence light was collimated using relay optics and input into themicroscope. In addition, for comparison, a highly-coherent source (633nm, Helium-Neon laser) was used in the input of the microscope.

In the microscope, a 40×, 0.66 numerical-aperture microscope objectiveMO and a 15 cm focal-length tube lens L₀ were used. The interferometer104, ported in the output of the microscope, contained two 7.5 cmfocal-length lenses L₁ and L₂, positioned in 4f configuration, a cubebeam splitter BS, and two mirrors M, with a pinhole P of 20 μmpositioned in front of one of them. The mirrors M were positioned veryclose to the output of the beam splitter, so that there was almost nopropagation through free space after splitting the beams and beforecombining them. No enclosure was used to avoid differential airperturbations between the interferometric arms. A monochrome digitalcamera (DCC1545M, Thorlabs) with 5.2 μm square pixels was positioned inthe output of the interferometer 104 to acquire the interferograms ofthe sample.

100 inteferograms per second were acquired and then processed into thephase profile of the sample by using a digital spatial filtering,followed by phase unwrapping algorithm for removing 2π ambiguities. Thefinal phase profile was obtained by subtracting the unwrapped phaseprofile from a sample-less interferogram, which compensates for(temporally-invariant) spatial noise. The resulting phase profile isproportional to the sample optical path delay profile.

FIG. 2 contains a plot of the unbiased optical path delay in nm againsttime in sec as obtained for the on-axis configuration. The dashed-linegraph 202 in FIG. 2 represents the temporal optical path delay that wasobtained using the interferometer 104 and the highly-coherent source fora representative diffraction-limited spot, with standard deviation of0.41 nm. For comparison, the dotted-line graph 201 in FIG. 2 representsthe temporal optical path delay that was obtained using a conventionalMichelson interferometer under the same conditions (without using anenclosure) for a representative diffraction-limited spot, with astandard deviation of 2.4 nm. As shown by the solid-line graph 203 inFIG. 2, when using the low-coherence source, the interferometer 104yielded temporal optical path delay for a representativediffraction-limited spot with standard deviation of 0.18 mm Underlow-coherence illumination, spatially averaging the optical path delayprofile for 100×100 diffraction-limited spots yielded a standarddeviation of 0.42 nm, whereas a Michelson interferometer using thehighly-coherent source yielded a spatial standard deviation of 3.8 nm,mostly due to the presence of speckle noise.

Referring to FIG. 3, the thickness profile of a live human red bloodcell obtained by a single exposure using the on-axis interferometer 104and the low-coherence source 101 is shown. To obtain this thicknessprofile, the optical path delay profile of the cell was divided by thedifference between the refractive index of the cell (n=1.395), under theassumption of homogenous refractive index for an enucleated red bloodcell, and the refractive index of the surrounding media (n=1.34). Asshown in FIG. 3, due to the use of a low-coherence source, thebackground around the red blood cell (containing only cell media) isremarkably flat, with a standard deviation of spatially-averaged opticalpath delay of 0.85 nm in liquid environment. The invention provides asimple, cost-effective technology for significantly reducing the size ofthe interferometer (as low as 1 inch) and increasing the interferometermeasurement stability and thus its accuracy by using efficientcommon-path geometry.

Reference is made to FIG. 4A, illustrating another embodiment of thepresent invention in which the novel interferometer has an off-axisgeometry capable of creating a full off-axis interference pattern on thecamera. In order to create a small angle between the sample beam and thereference beam, and enable an off-axis interferogram, the actual Fourierplane center, described by the continuation of the reflected beam BL inFIG. 4A, is shifted using a retro-reflector RR. This retro-reflector maycomprise a pair of mirrors attached to each other in a right angle. FIG.4B shows the RR operation in tilting the sample beam and creating anoff-axis interferometric angle on the camera. This figure presents thetwo beams ray tracing as it would be seen if they both were on the sameoptical axis (so that a beam splitter was not used in the middle of the4f device composed of lens L1 and L2, but still the splitting would beperformed).

As can be seen from this figure, the retro-reflector creates an angle θbetween the beams, which is described as follows:

θ=arctan(Δy/f),  (1)

where θ is the angle between the reference beam and the sample beam, Δyis the shift between the focal points of the two beams, and f is thefocal length of lens L2.

To demonstrate the operation of the off-axis interferometer of thepresent invention, the inventors have constructed the experimental setupillustrated in FIG. 5. FIG. 5 presents a device 300 configured accordingto some embodiments of the present invention, including aninterferometric device 304 which in the present not limiting example isincorporated in an microscope being ported into the microscope output(replacing a digital camera typically installed there in themicroscope). The microscope includes a light source 301, such as alow-coherence laser, a sample holder Sample, and a microscope objectiveMO. Also, the microscope may include a light directing optics, such as alight deflector (mirror) M that directs light 302 from the light sourceonto a sample, and a tube lens L_(o). The objective lens and the tubelens create an appropriately magnified image of the illuminated spot ofthe sample on an image plane located in the interferometric device 304.The solid lines are directly transmitted light while the dashed linesare the image forming beams. In order to demonstrate the capabilities ofthe invention, the inventor performed an experiment with a device thatis similar to the device illustrated in FIG. 5 with the followingspecifications: this setup contains a simple invert microscope with asingle 40×, 0.66-numerical-aperture, infinity-corrected microscopeobjective, spherical tube lens with 15 cm focal length, and amonochromatic CMOS camera with 5.2 μm square pixels (Thorlabs DCC1545M).The off-axis interferometer 300 is connected between the microscopecamera port and the digital camera in a 4f configuration. In thisnon-limiting example, the regular microscope is illuminated by a tunablelow-coherence source 301. The light source 301 used in the input of theinvert microscope is a supercontinuum fiber-laser source (SC400-4,Fianium), connected to a computer-controlled acousto-optical tunablefilter (SC-AOTF, Fianium), tuned to a central wavelength of 633 nm and afull-width-at-half-maximum bandwidth of 6.7 nm, as measured by a compactspectrometer (USB4000-VIS-NIR, Ocean Optics). To collimate the beam inthe output of the tunable filter and to increase its spatial coherence,the beam was spatially filtered using 10× and 5× microscope objectivesand 25 μm confocally-positioned pinhole, creating magnification of 0.5.Lenses L1 and L2 were chosen to be achromatic lenses with focal lengthsof 100 mm and 125 mm respectively. The total magnification of theexperimental setup was 47× and the experimentally-confirmeddiffraction-limited spot was 0.815 μm. The interference area on thecamera sensor was 5.32 mm×5.32 mm (1024×1024 pixels of 5.2 μm each) witha high-visibility modulation area (above half of the maximum visibility)of 2.672 mm×5.32 mm (512×1024 pixels) and a fringe frequency of 48 linesper mm (fringe cycle of 4 pixels).

Based on the Fraunhofer diffraction through a circular aperture and theused optical elements in the off-axis interferometer of the presentinvention, for the experimental setup in FIG. 5, the pinhole diameter ischosen to be 30 μm. This pinhole size ensures that most of thesample-image data is erased and that the first airy disk defined by thepinhole covers most of the camera sensor where the interference fringesappear [7]. In the experimental setup, the camera was set on the maximumexposure time possible without reaching saturation, gamma value of 1 andno gain.

In addition to its portability, simple and inexpensive design, one ofthe advantages of the off-axis interferometer of the present inventionis its simple alignment. Using this interferometer, obtaininginterference with a high-coherence source, such as a HeNe laser, isimmediate, and the alignment with a low-coherence source issignificantly easier compared to obtaining low-coherence interferencewith conventional interferometers such as Mach-Zehnder or Michelsoninterferometers. The alignment of the off-axis interferometer of thepresent invention using a low-coherence source is done by firstlyaligning the pinhole on the combined focal point of lenses L1 and L2 andobtaining a circular diffraction image on the camera plane. Followingthis, the retro-reflector RR is positioned in such a way that on both ofits mirrors, the beam spot has the same size, while both spots appear asclose as possible to the connection between the two mirrors. By doingso, the optical path delay between the two beams will be far onlyseveral millimeters from interference in an on-axis geometry. Then, RRis shifted in z direction until an interference pattern between thebeams occurs. Following this, shifting RR in the y direction creates anoff-axis interference pattern on the camera, with an angle determined byEq. (1).

Since the phase of the wave reflected from the pinhole still containsthe DC frequency of the original wave, caused by the constant opticalthickness of parts in the entire sample (such as a cover-slip), theoptical thickness in the first exponent numerator (and in the secondone) lacks this constant value. This increases the fringes visibility,reduces noise, and more importantly, prevents changes in theinterference area due to positioning of samples with differentcover-slip thicknesses.

The following are some experimental results obtained with the device ofthe invention:

To estimate the off-axis configuration, spatial and temporal noiselevels, which determine the optical-path-delay sensitivity across animage and between images, respectively, two different samples wererecorded, one of a plain cover-slip and a second one of a chambercontaining only water. For this experiment, 150 interferograms of512×512 camera pixels were continuously recorded during 10 seconds. FIG.6A presents the standard deviation distribution of theoptical-path-delay map of 512×512 pixels for 150 interferograms for thedry sample (each value is from a different interferogram). Thisdistribution represents the spatial sensitivity/stability of the deviceof the present invention indicative of the spatial noise in theoptical-path-delay maps. As can be seen, most of the values are around0.6 nm. In the center of the image (a central area of 150×150 pixels),where the visibility is higher than 0.75, the spatial stability valuewas only 0.35 nm. FIG. 6B shows the standard deviation distribution pera single diffraction-limited spot across the 150 optical-path-delay mapsfor the dry sample, representing the temporal sensitivity/stability ofthe device. The mean temporal stability measured was 0.5 nm, where inthe center of the image it was 0.24 nm. For the wet sample, the temporalsensitivity had a mean value of 0.54 nm, whereas in the center of theimage it was only 0.28 nm.

The digital phase extraction of the sample phase from the interferogramis carried out by digital spatial filtering of the off-axisinterferogram, which includes a digital two-dimensional Fouriertransform, separation of the G₊₁ temporal coherence function from thezero-order I_(s)+I_(r), and back Fourier transform of the centered G₊₁temporal coherence function. Then, the phase argument of the resultingcomplex function is taken to obtain the wrapped phase. Afterwards, tocompensate for aberrations and field curvatures, the same wrapped-phaseextraction process is performed for a sample-free interferogram, and theresult is subtracted from the first wrapped phase. Finally thequality-guided two-dimensional unwrapping algorithm is applied to remove2π ambiguities.

Under the assumption of a constant immersion medium thickness andrefractive index around the sample, the optical thickness oroptical-path-delay profile of the sample (OPD_(s)) can be extracted bysubtracting all the elements having a constant refractive index andthickness, and written as follows:

OPD_(s)(x,y)=[ n _(s)(x,y)−n _(m) ]×h _(s)(x,y),  (2)

where n_(m) is the constant refractive index of the immersion medium,h_(s) is the thickness profile of the sample, and n _(s) is the sampleintegral refractive index which is defined as follows:

$\begin{matrix}{{{\overset{\_}{n}}_{s}\left( {x,y} \right)} = {\frac{1}{h_{s}}{\int_{0}^{h_{s}}{{n_{s}\left( {x,y,z} \right)}\ {{z}.}}}}} & (3)\end{matrix}$

To assess the capabilities of the off-axis interferometer of the presentinvention, measurements on several targets were performed and comparedwith the performances of a modified Mach-Zehnder interferometer, acommon IPM setup [P. Girshovitz and N. T. Shaked, “Generalized cellmorphological parameters based on interferometric phase microscopy andtheir application to cell life cycle characterization,” Biomed. Opt.Express 3, 1757-1773 (2012).], when using both a high-coherence source(HeNe laser) and a low-coherence source with 6.7 nm spectral bandwidth.The comparative results are presented between the devices under theexact same conditions (where the devices operated using the samelow-coherence light source, camera, optical table, etc.) The same fringevisibility area was chosen and the same reconstruction algorithms wereapplied in all cases.

The first measured sample was a new 600 lp/mm volume phase holographicgrating (from Edmunds Optics). This grating is characterized by aconstant physical thickness and a periodic refractive index with aconstant amplitude and frequency. FIGS. 7A-7B are optical-path-delay oroptical thickness maps of a volume phase holographic grating obtainedunder low-coherence illumination by the off-axis interferometer of thepresent invention; and a Mach-Zehnder interferometer respectively. FIGS.7A-7B present the measurements done on the center of the grating usingboth the off-axis interferometer of the present invention and theMach-Zehnder interferometer, both using the same low-coherence source.By comparing the results of both setups, it can be seen that both setupsmanaged to recognize the periodic refractive index of the grating.However, the off-axis interferometer of the present invention provided asignificantly clearer and more consistent image (see FIG. 7A) comparedto the conventional Mach-Zehnder interferometer (see FIG. 7B), whichyielded artifacts like brakes in the ridges and inconstant base plane.

Using FIB lithography, the inventors created several custom-made phasetargets containing variable depths elements such as shapes on achrome-plated glass cover-slip (10 nm plating) in different heights,ranging from 10 nm to 300 nm. The first element was a large plate-likeshape with a curvature inside. Six smaller plate-like shapes werepositioned symmetrically inside the large plate with a deeper curvaturesand a logo was lithographed in the middle of the larger plate with adeeper milling as illustrated in FIG. 8.

Reference is made to FIGS. 9A-9C presenting optical-path-delay maps ofthe phase target of FIG. 8 as obtained by the off-axis interferometer ofthe present invention using a low-coherence source (FIG. 9A), by aMach-Zehnder interferometer using a low-coherence source (FIG. 9B), andby a Mach-Zehnder interferometer using a high-coherence source (HeNelaser) (FIG. 9C). While the three configurations managed to record thelogo in the center of the plate, the modified Mach-Zehnderinterferometer failed in recording the large plate curvature, as well ascould hardly visualized the smaller six plates around the logo, due tohigher spatial noise level, which is even severer in the coherent setup(FIG. 9C), as expected. In FIG. 9C, the effects of the coherent noiseand the self interferences of the high-coherence source distorted thethinnest elements, i.e. the six plates and larger plate.

A second phase target containing variable depths elements, lithographedby the same FIB technique, contained the words “OMNI Group” with a linewidth of 0.7 μm (close to the microscope diffraction-limit spot) and anoptical thickness of 20 nm (10 nm due to the milling of the chrome and10 nm due to the milling of the glass). Again, three cases werecompared: the off-axis τ interferometer using a low-coherence source, aMach-Zehnder interferometer using a low-coherence source, and aMach-Zehnder interferometer using a high-coherence source (HeNe laser).The corresponding optical-path-delay maps are shown in FIGS. 10A-10C. Asit was for the first target of FIG. 5, the lower spatial noise level ofthe off-axis interferometer of the present invention enables it to seesmaller features that the other conventional IPM setups cannot see. InFIG. 10A, the lithographed text “OMNI Group” is clearly seen anddistinguished from the background, whereas in the measurements done bythe modified Mach-Zehnder interferometer, presented in FIGS. 10B-10C,the background noise level conceals most of the lithographed text andonly several lines are barely seen.

The mean optical-path-delay of the lithographed text letters in FIG. 10Ais measured as 20 nm, which agrees with the real optical-path-delay ofthe letters that was calculated as 19.4 nm (n_(chrome)=2.42 andn_(glass)=1.515). It should be noted that minimal milling capability ofthe FIB setups used in the experiments is 10 nm, so it is possible thatthe inconstant optical-path-delay of the letters seen in FIG. 11A iscaused by the milling process of the glass layer and not due to thespatial interferometric noise. In any case, these results show that theoff-axis interferometer of the present invention can be used to performinexpensive quality checks and imaging during the manufacturing oftransparent optical elements, as long as the lateral dimensions of thesmallest element that need to be examined is larger than thediffraction-limit spot of the microscope.

The inventors have also measured red blood cell (RBC) membranefluctuations, where 300 frames at 25 frames per second were captured.FIGS. 11A-11D show the optical-path-delay and physical thickness profileof RBC sample from both setups while using a low-coherence source of theoff-axis interferometer of the present invention and of the off-axisMach-Zehnder interferometer (FIGS. 11A and 11B, respectively), and thecorresponding temporal standard deviation maps of the off-axisinterferometer of the present invention and of the off-axis Mach-Zehnderinterferometer (FIGS. 11C and 11D, respectively). The bar in the leftside of FIG. 11 is made to show both the optical-path-delay values andthe physical thickness values based on the refractive indices assumptionof 1.395 for the RBCs and 1.33 for the medium.

The optical-path-delay profiles show a slightly lower spatial noiselevel in the off-axis interferometer of the present invention (FIGS.11A-11B, with more self-interferences in the bottom right side of FIG.11B). It should be noted that none of the images have been digitallytreated to improve its quality. The standard deviation calculated byusing the Mach-Zehnder interferometer showed higher temporal noiselevels with a mean background value of 1.1 nm, compared to only 0.32 nmin the off-axis interferometer of the present invention (FIGS. 11C-11D).The standard deviations of the RBC optical-path-delay fluctuations weresimilar for the two types of measurements, ranging from 0.7 nm to 7 nm[I. Shock, A. Barbul, P. Girshovitz, U. Nevo, R. Korenstein, and N. T.Shaked, “Optical phase measurements in red blood cells usinglow-coherence spectroscopy,” J. Biomed. Opt. 17, 101509 (2012)]. Itshould be noted that not all of fluctuations were seen using theMach-Zehnder interferometer, as in some cases the temporal noise levelwas higher than the lowest measurable optical-path-delay standarddeviation.

One of the advantages of the off-axis interferometer over the on-axisinterferometer is the capability of recording dynamic changes in a largefield of view, where the frame rate is limited only by the maximal framerate of the camera sensor (since only one frame of acquisition isrequired to reconstruct the optical-path-delay map), and the field ofview is only limited by the complex degree of coherence of the lightsource used and not by the optical system. FIG. 12 presents Blepharismaorganism in motion using the off-axis interferometer of the presentinvention, as it swims through the entire field of view in water at aframe rate of 25 fps. The entire motion across the field of view lastedless than half a second, where there is not a point in time where theBlepharisma was stationery. As can be seen in FIG. 12, at 0 msec, 40msec, 320 msec and at 360 msec, due to the low-coherence length of thelight source, the reconstruction of the Blepharisma image is notcomplete. In the first two cases (0 and 40 msec), the lower coherencelength causes a low visibility of the interference fringes in some areasat the edges of the frame. In these areas, the interference visibilityis low and the phase cannot be well reconstructed. In the last two cases(320 and 360 msec), the Blepharisma is positioned in an angle to theimaging plane, which causes an erroneous reconstruction in theseout-of-focus points, so not all parts of the organism can bereconstructed. These problems may be solved by using a coherent sourceas the coherence length will increase, however coherent noise will behigher while decreasing the device sensitivity [7]. This figuredemonstrates the device capabilities for quantitative imaging of fastdynamics on relatively large field of view due to its true off-axisconfiguration.

In addition to the facts that the interferometer of the presentinvention is smaller, less expensive, more portable and significantlyeasier to construct and to align compared to the conventional off-axisIPM setups, the experimental results have shown that the off-axisinterferometer of the present invention provides better and cleareroptical-path-delay maps, with lower spatial and temporal noise.

The off-axis interferometer of the present invention is able to record asingle interferogram containing more than 1 Mega pixels (an area of 5.32mm×5.32 mm on our camera sensor), and due to its off-axis geometry, itallows multi-cells imaging in a single frame of acquisition. Thetemporal and spatial noises, determining the smallest dynamic change andthe smallest optical path delay that may be detected by the setup, arein the sub-nanometer range with values less than 0.7 nm for the fullimage and less than 0.4 nm in the center of the image. The off-axisinterferometer of the present invention have recorded objects withthicknesses of about 20 nm, which otherwise would be obscured by thespatial noise, as was demonstrated using FIB-lithographed elements, andhave detected dynamic changes in the range of 10 nm.

In some embodiments, the device may comprise a diffraction gratingconfigured for tilting the field in one of the beams to create afull-field, high-visibility interference on the entire camera plane [7].

1-23. (canceled)
 24. An interferometric device comprising a lightdirecting optical arrangement for directing light to an opticaldetector, wherein the light directing optical arrangement is configuredfor defining first and second substantially overlapping optical pathstowards said optical detector, the light directing optical arrangementcomprises: a beam splitter/combiner unit for receiving an input beam ofan amplitude and phase modulation and splitting said input beam intofirst and second light beams; a first and second reflective surfacesaccommodated in the first and second optical paths of said first andsecond light beams to direct said first and second light beams back tosaid beam splitter/combiner that directs the combined beam to thedetector; a spatial filter comprises a pinhole accommodated in front ofat one of the first and second reflective surfaces and a Fourier opticsassembly comprising two lenses, one lens being in a 4f configurationwith respect to the other; the beam splitter/combiner unit directing oneof the first and second light beams through said spatial filter toenable amplitude and phase demodulation thereof and formation of areference beam with respect to the other modulated beam.
 25. Theinterferometric device of claim 24, wherein said pinhole is located in apredetermined Fourier plane.
 26. The interferometric device of claim 24,wherein one of said first and second reflective surfaces comprises aretro-reflector.
 27. The interferometric device of claim 24, whereinsaid beam splitter/combiner unit comprises a cube beam splitter.
 28. Theinterferometric device of claim 24, wherein said first and secondreflective surfaces are placed at the outputs of the beamsplitter/combiner unit.
 29. The interferometric device of claim 24,wherein one of said two lenses is located at its focal length from thedetector.
 30. The interferometric device of claim 24, wherein one of thefirst and second reflective surfaces is located after one of the twolenses at a distance of the focal length of the lens.
 31. Theinterferometric device of claim 24, comprising a phase shifting deviceinto one of the beam paths.
 32. An optical system comprising: a beamsplitter/combiner unit for receiving an input beam of certain amplitudeand phase modulation and splitting said input beam into first and secondlight beams of the same amplitude and phase modulation and combiningreflections of the first and second light beams to produce an outputcombined beam; a first and second reflective surfaces accommodated inthe first and second optical paths of the first and second light beamsto thereby direct the first and second light beams back to the beamsplitter/combiner that directs the combined to the detector; a spatialfilter comprising a pinhole accommodated in front of one of the firstand second reflective surfaces in the optical path of the first splitlight beam to apply amplitude and phase demodulation thereto and therebyform a demodulated reference beam with respect to the second modulatedbeam and a Fourier optics assembly comprising two lenses, a second lensbeing positioned in an 4f configuration with the first lens; therebyenabling an interference pattern resulting from interaction of thereference and modulated beams to be indicative of said amplitude andphase modulation.
 33. The optical system of claim 32, wherein one ofsaid first and second reflective surfaces comprises a retro-reflector.34. The optical system of claim 32, wherein said beam splitter/combinerunit comprises a cube beam splitter.
 35. The optical system device ofclaim 32, wherein said first and second reflective surfaces are placedat the outputs of the beam splitter/combiner unit.
 36. The opticalsystem of claim 32, wherein one of said two lenses is located at itsfocal length from the detector.
 37. The optical system of claim 32,wherein one of the first and second reflective surfaces is located afterone of the two lenses at a distance of the focal length of the lens. 38.The optical system of claim 32, comprising a phase shifting device intoone of the beam paths.
 39. A sample inspection system, comprising: lightcollecting and focusing optics configured and operable for collecting aninput beam from a predetermined sample surface and focusing it onto animage plane; an interferometer unit accommodated in a path of the lightcollected by the light collecting and focusing optics, theintereferometer unit comprising: a beam splitter/combiner unit forreceiving the input beam of certain amplitude and phase modulation andsplitting said input beam into first and second light beams of the sameamplitude and phase modulation and combining reflections of the firstand second light beams to produce an output combined beam; a first andsecond reflective surface accommodated in the first and second opticalpaths of the first and second light beams to thereby direct the firstand second light beams back to the beam splitter/combiner that directsthe combined to the detector; a spatial filter comprising a pinholeaccommodated in front of one of the first and second reflective surfacesin the optical path of the first split light beam being located in aFourier plane with respect to said predetermined surface to therebyapply amplitude and phase demodulation thereto and form a demodulatedreference beam with respect to the second modulated beam and a Fourieroptics assembly comprising two lenses, a second lens being positioned inan 4f configuration with the first lens; an interference patternresulting from interaction of the reference and modulated beams in theimage plane being thereby indicative of said amplitude and phasemodulation.
 40. The sample inspection system of claim 39, wherein saidpinhole is located in a predetermined Fourier plane.
 41. The sampleinspection system of claim 39, wherein one of said first and secondreflective surfaces comprises a retro-reflector.
 42. The sampleinspection system of claim 39, wherein said beam splitter/combiner unitcomprises a cube beam splitter.
 43. The sample inspection system ofclaim 39, wherein said first and second reflective surfaces are placedin the outputs of the beam splitter/combiner unit.
 44. The sampleinspection system of claim 39, wherein one of said two lenses is locatedat its focal length from the detector.
 45. The sample inspection systemof claim 39, wherein one of the first and second reflective surfaces islocated after one of the two lenses at a distance of the focal length ofthe lens.
 46. The sample inspection system of claim 39, comprising aphase shifting device into one of the beam paths.